Diabetic nephropathy (DN) develops in approximately 30% of diabetic patients, and represents the leading cause of end-stage renal disease worldwide. The factors that induce DN in some but not all diabetics are poorly understood. Identification of novel pathogenic factors that serve as biomarkers for DN is needed. Semaphorin3a (sema3a), a guidance protein secreted by podocytes, is essential for normal glomerular filtration barrier development. Sema3a gain-of-function prevents slit-diaphragm development and leads to proteinuric glomerular disease in adult mice. We identified the signaling pathway that links sema3a signals to nephrin and the podocyte actin cytoskeleton mechanistically. We observed that podocyte sema3a is increased in human advanced DN. Consistently, circulating sema3a and urinary excretion are elevated in diabetic mice, and advanced DN exacerbates this abnormal sema3a excretion. Moreover, Sema3a gain-of- function induced severe DN. Collectively, these findings raise the possibility that elevated sema3a levels drive glomerular damage in the diabetic milieu, or accentuate such damage, making sema3a a suitable DN biomarker and a potential therapeutic target. The goals of this proposal are to investigate whether sema3a is a pathogenic determinant or a biomarker of diabetic nephropathy (or both), and to identify the molecular mechanisms involved, with the ultimate objective of developing a novel approach to treat or prevent DN. We hypothesize that diabetes-induced excess sema3a negatively regulates nephrin signaling and F-actin via a pathway that involves sema3a receptor plexinA1, downstream signaling, thereby disrupting slit-diaphragms, causing foot process effacement and proteinuria. Aim 1 will investigate whether sema3a is pathogenic in DN using a T1D model in mice with inducible Sema3a gain-of-function or loss-of-function, assess DN phenotype reversibility and whether sema3a inhibition improves DN; to evaluate whether sema3a is a DN biomarker, urine and plasma sema3a levels will be measured in T1D and T2D human cohorts by quantitative enzyme-linked immunoassay. Aim 2 will examine how the sema3a-plexinA1 pathway regulates podocyte signaling and F-actin dynamics by evaluating sema3a-induced nephrin turnover, plexinA1-MICAL interaction, MICAL requirement, and modulation of RhoA, CRMP and 3 integrin by sema3a. Results of the proposed experiments have the potential to establish sema3a as a novel biomarker of DN and to identify targets for therapeutic intervention in DN.